Aza cyclohexapeptide compounds

ABSTRACT

Ceretain aza cyclohexapeptide compounds have been found to have superior antibiotic properties. Novel processes for their preparation are also described.

This is a continuation of U.S. application Ser. No. 08/032,847, filedMar. 16, 1993, now U.S. Pat. No. 5,378,804, issued Jan. 3, 1995.

The present invention is directed to certain aza cyclohexapeptidecompounds and to processes for their preparation.

The aza cyclohexapeptide compounds of the present invention, Compound I(Seq ID Nos. 1-15) are characterized in having a nitrogen attached tothe cyclohexapeptide ting at the 5-carbon of the 4-hydroxy ornithinecomponent (hereinafter "C-5-orn") and may be represented by the formula##STR1## wherein R₁ is H or OH

R₂ is H, CH₃ or OH

R₃ is H, CH₃, CH₂ CN, CH₂ CH₂ NH₂ or CH₂ CONH₂

R^(I) is C₉ -C₂₁ alkyl, C₉ -C₂₁ alkenyl, C₁ -C₁₀ alkoxyphenyl or C₁ -C₁₀alkoxynaphthyl

R^(II) is H, C₁ -C₄ alkyl, C₃ -C₄ alkenyl, (CH₂)₂₋₄ OH, (CH₂)₂₋₄ NR^(IV)R^(V), CO(CH₂)₁₋₄ NH₂

R^(III) is H, C₁ -C₄ alkyl, C₃ -C₄ alkenyl, (CH₂)₂₋₄ OH, (CH₂)₂₋₄NR^(IV) R^(V), or

R^(II) and R^(III) taken together are --(CH₂)₄ --, --(CH₂)₅ --, --(CH₂)₂O(CH₂)₂ -- or --(CH₂)₂ --NH--(CH₂)₂ --

R^(IV) is H or C₁ -C₄ alkyl

R^(V) is H or C₁ -C₄ alkyl; and

acid addition salts thereof.

Where the expression "alkyl", "alkenyl" or "alkoxy" is employed, it isintended to include branched as well as straight chain radicals.

The compounds of the present invention are generally obtained asmixtures of stereoisomeric forms in which one form usually predominates.Conditions may be adjusted by means within the normal skill of theskilled artisan to obtain predominantly the desired isomer. Thecompounds with preferred stereoisomeric form designated herein as the"normal" form may be seen in the working examples with the dashed linesbelow the plane at the "C-5-orn" position. The designation "epi" hasbeen employed for those compounds in which the group at the "C-5-orn"position is above the plane.

Pharmaceutically acceptable salts suitable as acid addition salts arethose from acids such as hydrochloric, hydrobromic, phosphoric,sulfuric, maleic, citric, acetic, tartaric, succinic, oxalic, malic,glutamic and the like, and include other acids related to thepharmaceutically acceptable salts listed in Journal of PharmaceuticalScience, 66, 2 (1977).

Representative nuclei for the aza derivatives of the present invention(Compound I) and the sequence ID for these compounds may be seen in thefollowing table. Since the peptide nuclei would be the same irrespectiveof substituents R^(I), R^(II) or R^(III), and since the sequenceidentification number is assigned for the nuclear variations, the aminesand salts have the same sequence ID's.

    ______________________________________    Aza                                 SEQ ID    Compound  R.sub.1  R.sub.2 R.sub.3  NO    ______________________________________    I-1       H        H       CH.sub.2 CONH.sub.2                                        1    I-2       H        H       CH.sub.2 CN                                        2    I-3       H        H       CH.sub.2 CH.sub.2 NH.sub.2                                        3    I-4       OH       H       CH.sub.2 CONH.sub.2                                        4    I-5       OH       H       CH.sub.2 CN                                        5    I-6       OH       H       CH.sub.2 CH.sub.2 NH.sub.2                                        6    I-7       OH       CH.sub.3                               CH.sub.2 CONH.sub.2                                        7    I-8       OH       CH.sub.3                               CH.sub.2 CN                                        8    I-9       OH       CH.sub.3                               CH.sub.2 CH.sub.2 NH.sub.2                                        9     I-10     OH       CH.sub.3                               CH.sub.3 10     I-11     OH       CH.sub.3                               H        11     I-12     OH       OH      CH.sub.2 CONH.sub.2                                        12     I-13     OH       OH      CH.sub.2 CN                                        13     I-14     OH       OH      CH.sub.2 CH.sub.2 NH.sub.2                                        14     I-15     H        CH.sub.3                               CH.sub.3 15    ______________________________________

One of the compounds which is particularly outstanding for the controlof mycotic infections is a compound identifiable as Compound I-6 whereinR^(II) is H, R^(III) is CH₂ CH₂ NH₂ and R^(I) is 9,11-dimethyltridecyl(DMTD), and which may be referred to specifically as Compound I-6-1 (SeqID No. 6). ##STR2##

In the above designation I-6-1 refers to the first compound in which thenuclear arrangement is I-6. Since in all the compounds of the presentinvention the substituent at the "C-5-om" is nitrogen, the substituentson said nitrogen may vary and still all compounds which have the sameR₁, R₂ and R₃ would be Seq ID No. 6.

The compounds are soluble in lower alcohols, and polar aprotic solventssuch as dimethylformamide (DMF), dimethyl sulfoxide (DMSO) and pyridine.They are insoluble in solvents such as diethyl ether and acetonitrile.

The compounds of the present invention are useful as an antibiotic,especially as an antifungal agent or as an antiprotozoal agent. Asantifungal agents they are useful for the control of both filamentousfungi and yeasts. They are especially adaptable to be employed for thetreatment of mycotic infections in mammals, especially those caused byCandida species such as C. albicans, C. tropicalis and C.pseudotropicalis, Cryptococcus species such as C. neoformans andAspergillus species such as A. fumigatus, A. flavus, A. niger. They arealso useful for the treatment and/or prevention of Pneumocystis cariniipneumonia to which immune-compromised patients are especiallysusceptible as hereinafter described.

The compounds of the present invention may be prepared fromcyclopeptides having the formula ##STR3## by a series of reactions inwhich the oxygen atom at the "C-5-om" (which also may be referred to asthe hemiaminal position) is ultimately replaced by nitrogen. Thestarting materials may be natural products or modified natural productsas subsequently described. When R₁ is hydrogen instead of hydroxyl, theproduct aza compounds may be prepared by an alternate series ofreactions. The method applicable for the preparation of compounds inwhich R₁ may be either H or OH is first described.

The sequence IDs of the starting materials are seen in the followingtable:

    ______________________________________                                      Starting                                      Material    Compound R.sub.1 R.sub.2 R.sub.3  SEQ ID NO.    ______________________________________    A-1      H       H       CH.sub.2 CONH.sub.2                                      16    A-2      H       H       CH.sub.2 CN                                      17    A-3      H       H       CH.sub.2 CH.sub.2 NH.sub.2                                      18    A-4      OH      H       CH.sub.2 CONH.sub.2                                      19    A-5      OH      H       CH.sub.2 CN                                      20    A-6      OH      H       CH.sub.2 CH.sub.2 NH.sub.2                                      21    A-7      OH      CH.sub.3                             CH.sub.2 CONH.sub.2                                      22    A-8      OH      CH.sub.3                             CH.sub.2 CN                                      23    A-9      OH      CH.sub.3                             CH.sub.2 CH.sub.2 NH.sub.2                                      24     A-10    OH      CH.sub.3                             CH.sub.3 25     A-11    OH      CH.sub.3                             H        26     A-12    OH      OH      CH.sub.2 CONH.sub.2                                      27     A-13    OH      OH      CH.sub.2 CN                                      28     A-14    OH      OH      CH.sub.2 CH.sub.2 NH.sub.2                                      29     A-15    H       CH.sub.3                             CH.sub.3 30    ______________________________________

Compounds A-4 and A-7 have been identified in the literature (J.Antibiotics 45, 1855-60 Dec. 1992) as pneumocandin B_(o) andpneumocandin A_(o) when R^(I) =DMTD.

When in Compound A-1, R₁ and R₂ are represented by any of the possiblevariables and R₃ is --H, CH₃ or --CH₂ CONH₂ (Seq ID Nos. 16, 19, 22,25-27 and 30), they may be directly employed in the first method. WhenR₃ is --CH₂ CN or --CH₂ CH₂ NH₂, the group --CH₂ CONH₂ may be firstconvened to --CH₂ CN or --CH₂ CH₂ NH₂ as subsequently disclosed and allthe modified compounds (Seq ID Nos. 17-18, 20-21, 23-24, 28-29) used inthe first method, or alternatively, a compound in which R₃ is --CH₂CONH₂ may be employed to produce a compound with N at the hemiaminalposition, and the --CH₂ CONH₂ of the resulting product then converted to--CH₂ CN or --CH₂ CH₂ NH₂.

First, when R₁, R₂ and R₃ of the starting material are the same as thatin the product, the following sequence may be employed. ##STR4##

In Step A, the starting material Compound A (Seq ID Nos. 16-30),alkylthiol or arylthiol and acid are caused to react in an aproticsolvent under anhydrous conditions for time sufficient for reaction totake place with the formation of Compound B (Seq ID Nos. 31-45), seen inthe following table. Aminoethylthiol has been found to be useful forthis step.

    ______________________________________                                      Sulfur                                      Intermediate    Compound R.sub.1 R.sub.2 R.sub.3  SEQ ID    ______________________________________    B-1      H       H       CH.sub.2 CONH.sub.2                                      31    B-2      H       H       CH.sub.2 CN                                      32    B-3      H       H       CH.sub.2 CH.sub.2 NH.sub.2                                      33    B-4      OH      H       CH.sub.2 CONH.sub.2                                      34    B-5      OH      H       CH.sub.2 CN                                      35    B-6      OH      H       CH.sub.2 CH.sub.2 NH.sub.2                                      36    B-7      OH      CH.sub.3                             CH.sub.2 CONH.sub.2                                      37    B-8      OH      CH.sub.3                             CH.sub.2 CN                                      38    B-9      OH      CH.sub.3                             CH.sub.2 CH.sub.2 NH.sub.2                                      39     B-10    OH      CH.sub.3                             CH.sub.3 40     B-11    OH      CH.sub.3                             H        41     B-12    OH      OH      CH.sub.2 CONH.sub.2                                      42     B-13    OH      OH      CH.sub.2 CN                                      43     B-14    OH      OH      CH.sub.2 CH.sub.2 NH.sub.2                                      44     B-15    H       CH.sub.3                             CH.sub.3 45    ______________________________________

For Step A, suitable acids include strong organic acid and mineralacids. Examples of strong organic acids are camphorsulfonic acid,p-toluenesulfonic acid and methanesulfonic acid. Mineral acids includehydrochloric acid and hydrobromic acid. Camphorsulfonic acid ispreferred.

Suitable solvents include DMF, DMSO, 1-methyl-2-pyrrolidinone andhexamethyl phosphoric triamide (HMPA). DMF or DMSO is preferred.

The reaction is generally carried out at ambient temperature for from 1to about 10 days.

In carrying out the reaction, the cyclohexapeptide compound, the thiolcompound and acid are stirred together in a suitable solvent until thereaction is substantially complete. The reaction mixture then is dilutedwith water and flash chromatographed on reverse phase resins using 10 to40 percent acetonitrile/water (containing 0.1% trifluoroacetic acid) aseluant. Trifluoroacetic acid may hereinafter be designated "TFA". Thefractions containing the desired product may be concentrated andlyophilized and the lyophilized material purified by preparative highperformance liquid chromatography (HPLC).

Appropriate columns for HPLC are commercially available columns soldunder trade mark names or trade names such as "ZORBAX" (DuPont),"DeltaPak" (Waters), Bio-Rad (Bio-Rad), "LICHROPREP" RP18 (E. Merck).The specific columns are identified in the working examples.

In Step B, Compound C (Seq ID Nos. 31-45), a sulfone is obtained by theoxidation of Compound B. Suitable oxidizing agents or oxidants include"OXONE," (KHSO₅ ·KHSO₄ ·K₂ SO₄ 2:1:1, Aldrich Chemicals)metachloroperoxybenzoic acid, and peroxyacetic acid. The sequence ID ofCompound C is the same as that of Compound B since the atom attached tothe hemiaminal carbon is still sulfur. Thus, the sequence IDs of thesulfones are as follows:

    ______________________________________                                        Sulfone    Compound   R.sub.1 R.sub.2 R.sub.3  SEQ ID    ______________________________________    C-1        H       H       CH.sub.2 CONH.sub.2                                        31    C-2        H       H       CH.sub.2 CN                                        32    C-3        H       H       CH.sub.2 CH.sub.2 NH.sub.2                                        33    C-4        OH      H       CH.sub.2 CONH.sub.2                                        34    C-5        OH      H       CH.sub.2 CN                                        35    C-6        OH      H       CH.sub.2 CH.sub.2 NH.sub.2                                        36    C-7        OH      CH.sub.3                               CH.sub.2 CONH.sub.2                                        37    C-8        OH      CH.sub.3                               CH.sub.2 CN                                        38    C-9        OH      CH.sub.3                               CH.sub.2 CH.sub.2 NH.sub.2                                        39     C-10      OH      CH.sub.3                               CH.sub.3 40     C-11      OH      CH.sub.3                               H        41     C-12      OH      OH      CH.sub.2 CONH.sub.2                                        42     C-13      OH      OH      CH.sub.2 CN                                        43     C-14      OH      OH      CH.sub.2 CH.sub.2 NH.sub.2                                        44     C-15      H       CH.sub.3                               CH.sub.3 45    ______________________________________

The oxidation of the thioether (Compound B) to the sulfone (Compound C)is carried out with about two molar amounts of the oxidant. When onemolar amount of oxidant is employed, the product is a sulfoxide whichmay then be convened to the sulfone. The sulfoxides may be employed asan intermediate in the formation the aza compounds but the sulfone ispreferred. A slight excess over the two molar amount of the oxidizingagent is employed.

The reaction is carried out in an aqueous medium, preferably a mixtureof acetonitrile and water. About equal amounts are preferred although arange of 1:9 to 9:1 may be employed.

In carrying out the reaction, the oxidant is added to a solution ofCompound B (Seq ID Nos. 31-45) in 1:1 acetonitrile/water and the mixtureallowed to stand at ambient temperature for time sufficient to completethe reaction to obtain Compound C generally from about 30 minutes to onehour.

After completion of the reaction, the compound is recovered from thereaction mixture by diluting with water and chromatographing. Reversephase (C18) flash column chromatography is suitable in this purificationstep. The preferred eluting agent is 30-45 percent acetonitrile/water(0.1% TFA) in 5 percent step gradients. The appropriate fractions arelyophilized to recover the desired sulfone intermediate, Compound C (SeqID Nos. 31-45). The intermediate tends to be labile, thus the isolationshould be carried out as rapidly as possible.

Compound C may be converted to a compound having a nitrogen directlyattached to the "C-5-orn". As seen in the flow diagram, reaction ofCompound C with an alkali metal azide produces an azide at that position(Compound D) while reaction with an amine compound (ammonia or amine)produces an amino group at the "C-5orn" position, (Compound I). CompoundD is an important intermediate for most of the compounds of the presentinvention. Although Compound D has nitrogen at "C-5-orn", since it isnot a product, separate sequence ID Nos. are assigned for Compound D.Sequence ID Nos. for Compound D are found in the following table.

    ______________________________________                                        Azide    Compound   R.sub.1 R.sub.2 R.sub.3  SEQ ID    ______________________________________    D-1        H       H       CH.sub.2 CONH.sub.2                                        46    D-2        H       H       CH.sub.2 CN                                        47    D-3        H       H       CH.sub.2 CH.sub.2 NH.sub.2                                        48    D-4        OH      H       CH.sub.2 CONH.sub.2                                        49    D-5        OH      H       CH.sub.2 CN                                        50    D-6        OH      H       CH.sub.2 CH.sub.2 NH.sub.2                                        51    D-7        OH      CH.sub.3                               CH.sub.2 CONH.sub.2                                        52    D-8        OH      CH.sub.3                               CH.sub.2 CN                                        53    D-9        OH      CH.sub.3                               CH.sub.2 CH.sub.2 NH.sub.2                                        54     D-10      OH      CH.sub.3                               CH.sub.3 55     D-11      OH      CH.sub.3                               H        56     D-12      OH      OH      CH.sub.2 CONH.sub.2                                        57     D-13      OH      OH      CH.sub.2 CN                                        58     D-14      OH      OH      CH.sub.2 CH.sub.2 NH.sub.2                                        59     D-15      H       CH.sub.3                               CH.sub.3 60    ______________________________________

The azide may be obtained by adding alkali metal azide while stirring atambient temperature to a solution of the sulfone (Compound C; Seq. DNos. 31-45) in an aprotic solvent for time sufficient to complete thereaction with the formation of the azide as determined by HPLC analysis.The reaction mixture then may be diluted with aqueous acid such astrifluoroacetic acid and then chromatographed to separate the desiredazide (Compound D) from the reaction mixture. Reverse-phase (C18) flashcolumn chromatography using 10-25 percent acetonitrile/water (0.1% TFA)in 5 percent step gradients is suitable for this procedure.

The azide (Compound D) may then be reduced to a compound having a freeamino group which is among the products (Compound I, Seq ID Nos. 1-15)of the present invention.

The reduction may be carried out by mixing the azide compound (CompoundI) with Pd/C in a solvent such as glacial acetic acid and hydrogenatingunder balloon pressure for 10 to 20 hours. The product then may berecovered by first removing the catalyst by filtration and the tiltratelyophilized to obtain the amine compound (Seq ID 1-15) in which theamine is a primary amine.

The amine thus obtained may be convened into a substituted amine assubsequently described.

Compound I in which --NR^(II) R^(III) is represented by --NHCH₂ CH₂ NH₂or generically by --NH(CH₂)₂₋₄ NR^(IV) R^(V) may be prepared from thesulfone by a method in which a diamine H₂ N(CH₂)₂₋₄ NR^(IV) R^(V) iscaused to react with the sulfone (Compound C, Seq ID Nos. 31-45).

The reaction is carded out in an aprotic solvent such as thosepreviously named and at ambient temperature. About tenfold molar excessof the amine compound is employed. The reaction may be carded out overone to several hours.

In carrying out the reaction, the appropriate amine is added to asolution of the sulfone in anhydrous aprotic solvent and the reactionmixture stirred at ambient temperature to obtain Compound I (Seq ID Nos.1-15) in which the substituent at "C-5-orn" is --NR^(II) R^(III). Thedesired compound may then be recovered by diluting with aqueoustrifluoroacetic acid and then chromatographing. Reverse phase (C18)flash column chromatography eluting with 10 to 25% acetonitrile/water(0.1% TFA) in 5 percent step gradients is suitable. The appropriatefractions may be lyophilized to recover the product as atrifluoroacetate salt.

The trifluoroacetate salt may be convened by dissolving the salt inwater and passing through a Bio-Rad AG2-XS(Cl--) polyprep column andrecovering the product as the hydrochloride salt.

When R₁ in formula (I) is hydrogen, Compound I' (Seq ID Nos. 1-3, 15),the nitrogen may be introduced directly into the hemiaminal position bya reaction to form the azide, which then is reduced to an amine whichoptionally may be alkylated or acylated to obtain the ultimate product.The reaction is seen by the following flow diagram. ##STR5##

Although R¹ is hydrogen in some natural product cyclohexapeptides, R¹ ismore commonly hydroxyl. Thus, for a number of the compounds, Compound A'in the flow diagram is prepared as a first step from the correspondingcompound in which R¹ is OH.

The preparation of the reduced compound may be carried out by stirringthe appropriate hydroxy compound in LiClO₄ -diethyl ether at roomtemperature, adding trifluoroacetic acid, followed by triethylsilane andsubjecting the mixture to rapid stirring for from 4 to 10 hours or untilthe starting hydroxy compound is no longer detectable by analyticalHPLC. The reaction mixture is then poured into distilled water to obtainthe reduced product as precipitate which then is recovered byconventional procedures. The reduced product thus obtained may be usedwith or without purification in the preparation of the azide.

Products in which R₁ is H, may be obtained by adding the modifiedcyclohexapeptide to a preformed solution of HN₃. HN₃ may be preparedfrom sodium azide and trifluoroacetic acid. The reaction is allowed totake place at room temperature to obtain the azide product which may berecovered by conventional procedures and purified by HPLC.

The purified azide compound may be reduced to the amine compound byhydrogenating with palladium/carbon in a manner similar to thatpreviously described.

The amines, prepared as above and having a primary amino group --NH₂described, may then be alkylated by conventional means to obtain asubstituted amino group. Briefly, alkylation may be carried out bycausing an appropriately substituted alkyl halide to react with theamine (Compound I, NR^(II) R^(III) =NH₂ ; Sequence ID Nos 1-15) in anaprotic solvent in the presence of a base to obtain the monosubstitutedamine (Compound I, NR^(II) R^(III) =NHR^(II) wherein R^(II) is C₁ -C₄alkyl, C₃ -C₄ alkenyl, (CH₂)₂₋₄ OH, and (CH₂)₂₋₄ NR^(IV) R^(V)). Thelatter may be recovered from the reaction mixture by conventionalprocedures.

The amines, prepared as above described and having a primary amino group--NH₂, may be acylated by conventional means to obtain an acylated aminogroup. The acyl group contemplated is CO(CH₂)₁₋₄ NH₂. Since this is aprimary amino group, the amino of the acylating acid is protected suchas with a benzyloxycarbonyl group before the acylation is carded out. Anactivated ester such as the pentafluorophenyl ester is preferably used.The acylation may be carded out in an aprotic solvent in the presence ofbase such as diisopropylethylamine at ambient temperature for from oneto several hours to obtain the acylation product. The product may berecovered by diluting the reaction mixture with methanol and purifyingby HPLC. The protecting group may be removed by conventionalhydrogenolysis. (Compound I, --NR^(II) R^(III) =--NHCO(CH₂)₁₋₄ NH₂).

The amine compounds in which the amino group at the hemiaminal positionis totally substituted, i.e. when neither R^(II) nor R^(III) is ##STR6##hydrogen, are preferably prepared by reacting the sulfone (Compound BSeq ID No. 31-45) with an appropriately substituted amine R^(II) R^(III)NH. The reaction may be carried out by adding the amine to a stirredsolution of the sulfone for time sufficient for reaction to take place.The product may be recovered by purifying by preparative HPLC andlyophilizing the appropriate components.

The invention also embraces acid addition salts. The compound in thenormal course of isolation is obtained as an acid addition salt.Generally, it is as a trifluoroacetic acid salt. The salt thus obtainedmay be dissolved in water and passed through an anion exchange columnbeating the desired anion. The eluate containing the desired salt may beconcentrated to recover the salt as a solid product.

The compounds of the present invention are active against many fungi andparticularly against Candida species. The antifungal properties may beillustrated with the minimum fungicidal concentration (MFC)determination against certain Candida organisms in a microbroth dilutionassay carded out in a Yeast Nitrogen Base (DIFCO) medium with 1%dextrose (YNBD).

In a representative assay, compounds were solubilized in 100% dimethylsulfoxide (DMSO) at an initial concentration of 5 mg/ml. Once dissolved,the drug stock was brought to a concentration of 512 μg/ml by dilutionin water such that the final DMSO concentration was about 10 percent.The solution was then dispensed via a multichannel pipetter into thefirst column of a 96-well plate (each well containing 0.075 ml of YNBD),resulting in a drug concentration of 256 μg/ml. Compounds in the firstcolumn were diluted 2-fold across the rows yielding final drugconcentration ranging from 256 μg/ml to 0.12 μg/ml.

Four-hour broth cultures of organisms to be tested were adjusted using aspectrophotometer at 600 nm to equal a 0.5 McFarland Standard. Thissuspension was diluted 1:100 in YNBD to yield a cell concentration of1-5×10⁴ colony forming units (CFU)/ml. Aliquots of the suspension (0.075ml) were inoculated into each well of the microtiter plate resulting ina final cell inoculum of 5-25×10³. CFU/ml and final drug concentrationsranging from 128 μg/ml to 0.06 μg/ml. Each assay includes one row fordrug-free control wells and one row for cell-free control wells.

After 24 hours of incubation, the microtiter plates were shaken gentlyon a shaker to resuspend the cells. The MIC-2000 inoculator was used totransfer a 1.5 microliter sample from each well of the 96-wellmicrotiter plate to a single reservoir inoculum plate containingSabouraud dextrose agar (SDA). The inoculated SDA plates were incubatedfor 24 hours at 35° C. The results were as follows:

    __________________________________________________________________________                            ORGANISM    COMPOUND†        C. albicans    C.parapsilosis                                                  C. tropicalis    R.sub.1         R.sub.2           R.sub.3 R.sup.II, R.sup.III                            MY 1055                                 MY 1028                                      MY 1750                                           MY 1010                                                  MY 1012    __________________________________________________________________________    1)      H  H --CH.sub.2 CH.sub.2 NH.sub.2                   H; CH.sub.2 CH.sub.2 NH.sub.2                            0.250                                 0.125                                      0.125                                           0.125  0.125    2)      H  H --CH.sub.2 CONH.sub.2                   H; CH.sub.2 CH.sub.2 NH.sub.2                            1.000                                 0.500                                      1.000                                           1.000  0.500    3)      H  H --CH.sub.2 CH.sub.2 NH.sub.2                   H; H     0.125                                 <0.060                                      0.125                                           <0.060 0.060    4)      OH H --CH.sub.2 CH.sub.2 NH.sub.2                   H; CH.sub.2 CH.sub.2 NH.sub.2                            <0.060                                 0.125                                      <0.060                                           <0.060 <0.060    __________________________________________________________________________     *R.sup.I = DMTD;     †as acid addition salts

The compounds also show in vivo effectiveness against fungi which may bedemonstrated with the same compounds of the in vitro assay.

Growth from an overnight SDA culture of Candida albicans MY 1055 wassuspended in sterile saline and the cell concentration determined byhemacytometer count and the cell suspension adjusted to 3.75×10⁵cells/mi. Then 0.2 milliliter of this suspension was administered I.V.in the tail vein of mice so that the final inoculum was 7.5×10⁴cells/mouse.

The assay then was carded out by administering aqueous solutions ofCompound I at various concentrations intraperitoneally (I.P.), twicedaily (b.i.d.) for four consecutive days to 18 to 20 gram female DBA/2mice, which previously had been infected with Candida albicans in themanner described above. Distilled water was administered I.P. to C.albicans challenged mice as controls. After seven days, the mice weresacrificed by carbon dioxide gas, paired kidneys were removedaseptically and placed in sterile polyethylene bags containing 5milliliters of sterile saline. The kidneys were homogenized in the bags,serially diluted in sterile saline and aliquots spread on the surface ofSDA plates. The plates-were incubated at 35° C. for 48 hours and yeastcolonies were enumerated for determination of colony forming units (CFU)per gram of kidneys. Compounds (1), (2), (3) and (4) gave >99 percentreduction of recoverable Candida CFUs at 0.09 and 0.375 mg/kg I.P. twicedaily for four consecutive days.

The compounds of the present invention are also useful for inhibiting oralleviating Pneumocystis carini infections in immune-compromisedpatients. The efficacy of the compounds of the present invention fortherapeutic or anti-infection purposes may be demonstrated in studies onimmunosuppressed rats.

In a representative study, the effectiveness of Compound I-6-1 (R₁ =OH;R₂ =H; R₃ =CH₂ CH₂ NH₂ ; R^(I) =DMTD; R^(II) =H; R^(III) =CH₂ CH₂ NH₂)was determined. Sprague-Dawley rats (weighing approximately 250 grams)were immunosuppressed with dexasone in the drinking water (2.0 mg/L) andmaintained on a low protein diet for seven weeks to induce thedevelopment of pneumocystis pneumonia from a latent infection. Beforedrag treatment, two rats were sacrificed to confirm the presence ofPneumocystis carinii pneumonia (PCP); both rats were found to haveinfections. Five rots (weighing approximately 150 grams) were injectedtwice daily for four days subcutaneously (sc) with Compound I-6-1 in0.25 ml of vehicle (distilled water). A vehicle control was also carriedout. All animals continued to receive dexasone in the drinking water andlow protein diet during the treatment period. At the completion of thetreatment, all animals were sacrificed, the lungs were removed andprocessed, and the extent of disease determined by microscopic analysisof stained slides. The results of this study showed Compound I-6-1reduced P. carinii cysts in 5 rats by at least 90 percent when dosed at0.075 mg/kg with all rats surviving.

The outstanding properties are most effectively utilized when thecompound is formulated into novel pharmaceutical compositions with apharmaceutically acceptable carrier according to the conventionalpharmaceutical compounding techniques.

The novel compositions contain at least a therapeutic antifungal orantipneumocystis amount of the active compound. Generally, thecomposition contains at least 1% by weight of Compound I. Concentratecompositions suitable for dilutions prior to use may contain 90% or moreby weight. The compositions include compositions suitable for oral,topical, parenteral (including intraperitoneal, subcutaneous,intramuscular, and intravenous), nasal, and suppository administration,or insufation. The compositions may be prepacked by intimately mixingCompound I with the components suitable for the medium desired.

Compositions formulated for oral administration may be a liquidcomposition or a solid composition. For liquid preparation, thetherapeutic agent may be formulated with liquid carriers such as water,glycols, oils, alcohols, and the like; and for solid preparations suchas capsules and tablets, with solid carders such as starches, sugars,kaolin, ethyl cellulose, calcium and sodium carbonate, calciumphosphate, kaolin, talc, lactose, generally with lubricant such ascalcium stearate, together with binders disintegrating agents and thelike. Because of their ease in administration, tablets and capsulesrepresent the most advantageous oral dosage form. It is especiallyadvantageous to formulate the compositions in unit dosage form (ashereinafter defined) for ease of administration and uniformity ofdosage. Compositions in unit dosage form constitute an aspect of thepresent invention.

Compositions may be formulated for injection and may take such forms assuspensions, solutions or emulsions in oily or aqueous vehicles such as0.85 percent sodium chloride or 5 percent dextrose in water and maycontain formulating agents such as suspending, stabilizing and/ordispersing agents. Buffering agents as well as additives such as salineor glucose may be added to make the solutions isotonic. The compound mayalso be solubilized in alcohol/propylene glycol or polyethylene glycolfor drip intravenous administration. These compositions also may bepresented in unit dosage form in ampoules or in multidose containers,preferable with added preservative. Alternatively, the activeingredients may be in powder form for reconstituting with a suitablevehicle prior to administration.

The term "unit dosage form" as used in the specification and claimsrefers to physically discrete units, each unit containing apredetermined quantity of active ingredient calculated to produce thedesired therapeutic effect in association with the pharmaceuticalcarrier. Examples of such unit dosage forms are tablets, capsules,pills, powder packets, wafers, measured units in ampoules or inmultidose containers and the like. A trait dosage of the presentinvention will generally contain from 100 to 200 milligrams of one ofthe compounds.

When the compound is for antifungal use any method of administration maybe employed. For treating mycotic infections, oral or intravenousadministration is usually employed.

When the compound is to be employed for control of pneumocystisinfections it is desirable to directly treat lung and bronchi. For thisreason inhalation methods are preferred. For administration byinhalation, the compounds of the present inventions are convenientlydelivered in the form of an aerosol spray presentation from pressurizedpacks or nebulisers. The preferred delivery system for inhalation is ametered dose inhalation (MDI) aerosol, which may be formulated as asuspension or solution of Compound I in suitable propellants, such asfluorocarbons or hydrocarbons.

Although the compounds of the present invention may be employed astablets, capsules, topical compositions, insufflation powders,suppositories and the like, the solubility of the compounds of thepresent invention in water and aqueous media render them adaptable foruse in injectible formulations and also in liquid compositions suitablefor aerosol sprays.

The following examples illustrate the invention but are not to beconstrued as limiting.

Examples 1-3 illustrate the preparation of the products by is the firstmethod described, namely proceeding through the sulfone. This method maybe employed in the preparation of any of the compounds but must beemployed to obtain a useful yield of product when R₁ is OH.

Examples 4 and following illustrate preparation of the products bydirect substitution of nitrogen for oxygen into the hemiaminal position"5-orn". This method is preferred when R₁ is H, and R^(II) and R^(III)are H.

Example 3 illustrates employing as starting material, a compound inwhich R₃ has already been reduced to CH₂ CH₂ NH₂ from the naturalproduct state where R₃ is CH₂ CONH₂. Similarly for compounds in which R₃is --CH₂ CN, the already partially modified compound may be employed.

Examples 9 and 10 illustrate carrying out the conversion of thehemiaminal oxygen to nitrogen and then converting the CH₂ CN or CH₂ CH₂NH₂.

EXAMPLE 1 ##STR7##

Part A. Preparation of Intermediate 1-4-hydroxy-5-(epi)-aminoethylthio-N²-(10,12-dimethyl-1-oxotetradecyl)ornithine!-5-(3-hydroxyglutamine)-6-(3-hydroxyproline)echinocandinB (Seq ID No 34)

A solution of 500 mg (0.47 mmol) of pneumocandin B_(o) (Seq ID No 19),5.34g (47 mmol) of 2-amino-ethanethiol hydrochloride and 109 mg (0.47mmol) of (1S)-(+)-10-camphorsulfonic acid in 40 ml anhydrous DMF wasstirred at 25° C. for 6 days. The reaction mixture was diluted with 40ml of water and flash chromatographed on "LICHROPREP" RP18 (40-63 μm,15.0g) packed with 10% acetonitrile/water. The column was eluted with 10to 40% acetronitrile/water, collecting two 120 ml fractions at each 10percent gradient. From the two 40% acetonitrile/water fractions wasobtained 185 mg of material which was purified by preparative HPLC"ZORBAX" C8 (21.2×250 mm), eluting with 40-45% acetonitrile/water (0.1%TFA) to obtain 128 mg of 1- 4-hydroxy-5-(epi)-aminoethylthio-N²-(10,12-dimethylo-1-oxotetradecyl)-omithine!-5-(3-hydroxyglutamine)-6-(3-hydroxyproline)-echinocandinB trifluoroacetate as a white amorphous solid. ¹ H NMR (400 MHz, CD₃ OD)δ1.34 (d, J=6.3 Hz, 3H), 2.89 (m, 2H), 4.72 (d, J=4.9 Hz, 1H) FAB-MS(Li), m/e 1131 (MH+Li)⁺

Pan B. Preparation of Intermediate Sulfone (Seq. ID 34)

To a stirred solution of the thio compound (444 mg, 0.358 mmol) obtainedin Part A, in 15 mL of 1:1 acetonitrile/water was added "OXONE" (324 mgequivalent to 1.06 mmol of potassium hydrogen persulfate). After about45 minutes, the solution was diluted with an equal volume of water andrapidly chromatographed using reverse-phase (C18) flash chromatographycolumn eluting with 35-43% acetonitrile/water (0.1% TFA) in 2% stepgradients. The product containing fractions were lyophilized to obtain357 mg (86% yield) of the epi-sulfone. ¹ H NMR (400MHz, CD₃ OD) δ3.48(m, 2H), 3.55 (m, 1H), 3.71 (m, 1H), 3.91 (dd, 1H), 4.00 (m, 1H), 5.17(dd, 1H), 6.76 (d, 2H), 7.16 (d, 2H)

Part C. Preparation of Product of Formula (1); Compound I-4 (Seq ID No4)

To a stirred solution of 1.2 g (0.945 mmol) of epi-sulfone (prepared asdescribed in Pan B) in 20 mL of anhydrous DMF was added ethylenediamine(568 mg, 9.45 mmol). After 1 hour, HPLC analysis (RP-C18, 40% CH₃ CN/H₂O (0.1% TFA))of the reaction mixture indicated complete conversion totwo polar products in a ratio of 37:63. Reverse phase (C18) flash columnchromatography eluting with 10-40% acetonitrile/water (0.1% TFA) in 5percent step gradients was followed by lyophilization of the appropriatefractions to provide 200 mg (21% yield) of the normal product as the(bis)-trifluoroacetate salt. ¹ H NMR (400 MHz, CD₃ OD) δ1.14 (d, J=6.2Hz, 3H), 2.72 (dd, J=15.4 and 3.8 Hz, 1H), 4.10 (m, H), 5.04 (dd, J=8.7and 3.2 Hz, 1H), 5.09 (dd, J=8.5 and 4.2 Hz, 1H), 5.18 (br s, 1H), 6.74(d, J=8.6 Hz, 2H), 7.12 (d, J=8.6 Hz, 2H), 7.47 (d, J=8.6 Hz, 1H), 7.71(d, J=10.0 Hz, 1H), 8.11 (d, J=8.7 Hz, 1H), 8.71 (d, J=8.7 Hz, 1H).FAB-MS (Li), m/z 1113.5 (MLi)⁺

The (bis)-trifluoroacetate salt from above was dissolved in H₂ O and thesolution passed through a Bio-Rad AG2-X8 (Cl --) polyprep column washingwith additional water. The product-containing eluate was lyophilized togive the above compounds as the (bis)-hydrochloride salt. Lyophilizationof the fractions containing the major product provided epi-product ¹ HNMR (400 MHz, CD₃ OD) δ3.02 (m, 1H), 3.14 (m, 3H), 4.16 (m, 1H), 5.10(dd, 1H), 6.76 (d, 2H), 7.14 (d, 2H). FAB-MS (Li), m/z 1113.9 (MLi)⁺

EXAMPLE 2 ##STR8##

Part A. Preparation of Intermediate Sulfone (Seq. ID No. 36)

The starting compound, Compound A-6 R^(I) =DMTD (Seq. ID No. 21), wasprepared as described for such compound in the section entitledPreparation of Starting Materials.

Compound A-6 was then converted to the epi-thio compound Compound B-6(Seq ID. No. 36) in a manner similar to that as described in Pan A ofExample 1.

To a stirred solution of 285 mg (0.241 mmol) of Compound B-6 in 14 mL of1:1 acentonitrile/water was added "OXONE" (162 mg equivalent to 0.530mmol of potassium hydrogen persulfate). After a period of 45 minutes,the solution was diluted with an equal volume of water andchromatographed. Reverse-phase (C18) flash column chromatography elutingwith 30-45% acetonitrile/water (0.1% trifluoroacetic acid) in 5% stepgradients was followed by lyophilization of the product-containingfractions to provide 212 mg of the epi-sulfone (Compound C-6 Seq ID. No.36) Yield=84%. ¹ H NMR (400 MHz, CD₃ OD) δ3.08 (M, 2H), 3.46 (t, J=6.6Hz, 2H), 3.68 (m), 5.05 (M), 6.77 (d, J=8.5 Hz, 2H), 7.15 (d, J=8.5 Hz,2H) FAB-MS (Li), m/z 1039.9

Part B. Preparation of the Product of Formula (2)

(Compound I-6; R^(II) =R^(III) =2-aminoethyl); .Seq ID No. 6

To a stirred solution of Compound C-6 (prepared as described in Part A,418 mg, 0.305 mmol) in 10 mL of anhydrous N,N-dimethylformamide wasadded ethylenediamine (183 mg, 3.05 mmol). After a period of 1 h, HPLCanalysis (RP-C18, 35% CH₃ CN/H₂ O (0.1% CF₃ COOH)) of the reactionmixture indicated complete conversion to two polar products in a ratioof 36:64. The reaction mixture was diluted with aqueous trifluoroaceticacid (190 mL H₂ O, 0.4 mL CF₃ COOH) and chromatographed. Reverse-phase(C18) flash column chromatography eluting with 10-25% acetonitrile/water(0.1% trifluoroacetic acid) in 5% step gradients was followed bylyophilization of the appropriate fractions to provide 111 mg of theproduct as the (tris)-trifluoroacetate salt: Yield=25% ¹ H NMR (400 MHz,CD₃ OD) δ1.17 (d, J=6.2 Hz), 2.44 (dd, J=7.0 and 13.2 Hz, 1H), 2.7-3.0(m, 4H), 3.06 (t, J=7.0 Hz, 2H), 3.82 (m, 3H), 3.97 (dd, J=11.2 and 3.2Hz, 1H), 4.03 (m, 2H), 4.70 (d, J=2.3 Hz, 1H), 5.00 (d, J=3.3 Hz, 1H),6.75 Hz (d, J=8.6 Hz, 2H), 7.11 (d, J=8.6 Hz, 2H) FAB-MS (Li), m/z1099.9 (MLi)⁺, 1033.9

The (tris)-trifluoroacetate salt from above was dissolved in H₂ O andthe solution passed through a Bio-Rad AG2-X8 (Cl--) polyprep columnwashing with additional water. The product-containing eluate waslyophilized to give 93 mg of the above compound as the(tris)-hydrochloride.

EXAMPLE 3 ##STR9##

Part A: Preparation of Azide (Seq. ID No, 49)

To a stirred solution of 297 mg, 0.257 mmol epi-sulfone (Example 1, PanB) in 10 milliliters of anhydrous dimethylformamide was added lithiumazide (126 mg, 257 mmol). After a period of 1 hr, HPLC analysis (RP-18,40% CH₃ CN/H₂ O (0.1% of CF₃ COOH)) of the reaction mixture indicatedcomplete conversion to a single substantially less polar product.Reverse phase (C18) flash column chromatography eluting with 30-65%acetonitrile/water in 5% step gradients was followed by lyophilizationof the product-containing fractions to provide crude azide. PreparativeHPLC (C18, 40-45% CH₃ CN/H₂ O (0.1% CF₃ COOH) in one 5% step gradient)produced an azido compound, Compound D-4, (Seq. ID No. 49). ¹ H NMR (400MHz, CD₃ OD) δ1.14 (d, J=6.1 Hz, 3H), 2.50 (dd, J=15.6 and 9.9 Hz, 1H),2.84 (dd, J=15.6 and 3.3 Hz, 1H), 3.95 (dd, J=11.2 and 3.1 Hz, 1H), 4.05(m, 2H), 4.56 (m, 3H), 4.98 (dd, J=8.5 and 3.5 Hz, 1H), 5.10 (dd, J=8.3and 4.2 Hz, 1H), 5.26 (dd, J=8.5 and 2.2 Hz, 1H), 6.74 (d, J=8.6 Hz,2H), 7.12 (d, J=8.6 Hz, 2H), 7.44 (d, J=8.3 Hz, 1H), 7.76 (d, J=9.9 Hz,1H), 8.26 (d, J=8.1 Hz, 1H), 8.83 (d, J=8.7 Hz, 1H), 9.00 (d, J=8.5 Hz,1 H) FAB-MS (Li), m/z 1096.9 (MH+Li)⁺ IR (Nujol mull, cm⁻¹) 2110

Part B: Preparation of the Amine (Seq. ID No. 4)

A mixture of azido compound D-4, prepared in Part A, (137 mg, 0.126mmol) and 10% Pd/C (137 mg) in glacial acetic acid (10 mL) washydrogenated under balloon pressure for a period of 14 h. The catalystwas removed by filtration and the tiltrate was lyophilized to obtain thecrude amine. Purification by preparative HPLC (C 18, 35-41% CH₃ CN/H₂ O(0.1% CF₃ COOH) in 3% step gradients), followed by lyophilization of theappropriate fractions provided the aza compound, Compound I-1, R^(II),R^(III) =H (Seq. ID No. 1) as the trifluoroacetate salt: Yield=48% ¹ HNMR (400 MHz, CD₃ OD) δ1.13 (d, J=6.1 Hz, 3H), 2.49 (dd, J=15.6 and 9.8Hz, 1H), 2.81 (dd, J=15.6 and 3.4 Hz, 1H), 3.97 (dd, J=11.1 and 3.1 Hz,1H), 4.03 (m, 1H), 4.11 (m, 1H), 4.47 (dd, J=11.7 and 5.5 Hz, 1H), 4.57(m, 2H), 5.00 (m, 1H), 5.10 (m, 1H), 5.14 (d, J=2.2 Hz, 1H), 6.74 (d,J=8.6 Hz, 2H), 7.12 (d, J=8.6 Hz, 2H), 7.42 (d, J=8.3 Hz, 1H), 8.89 (d,J=8.8 Hz, 1H) FAB-MS(Li), m/z 1071.0 (MLi)⁺

The trifluoroacetate was dissolved in H₂ O and the solution passedthrough a Bio-Rad AG2-X8 (Cl--) polyprep column, washing with additionalwater. The product-containing eluate was lyophilized to obtain 66 mg ofcompound 14, R^(II), R^(III) =H (Seq ID No. 1) as the hydrochloride.

In the following experiments, Solvent A=95% water/5% acetonitrile/0.1%trifluoroacetic acid and Solvent B=95% acetonitrile/5% water/0.1%trifluoroacetic acid. When the expression "in vacuo" or "rotovaped" isused, it refers to removal of solvent on a rotary evaporator.

EXAMPLE 4 ##STR10##

A. Preparation of Intermediate Azide Compound D-1 (Seq ID No. 46)

Pneumocandin B₀ (Compound A-4; Seq ID No. 19) (5.00 g, 4.69 mmol) wasdissolved in 2M LiClO₄ -diethyl ether at room temperature.Trifluoroacetic acid (2.50 ml) was added to the stirring solutionfollowed by triethylsilane (5.00 ml). The heterogeneous mixture wasstirred rapidly for 6 hours after which time little or no startingpneumocandin B₀ was detectable by analytical HPLC (C18 "ZORBAX", 45%Solvent A/55% Solvent B/0.1% TFA, 1.5 ml/min). The mixture was pouredinto 200 ml of distilled water, filtered and air dried. The wet solidwas stirred with diethyl ether, filtered and air dried to obtain 5.6 gof crude monoreduced pneumocandin B₀. (Compound A-1; Seq ID No. 16).

The crude isolate from above was added, as a solid, to a preformedsolution of HN₃ prepared by dissolving NaN₃ (3.06 g, 47.0 mmol) in 100ml of trifluoroacetic acid with cooling. After stirring at roomtemperature for 30 minutes, the reaction mixture was poured into 350 mlof distilled water and stirred for 15 minutes. The precipitate wasfiltered, dissolved in methanol and the solvent removed in vacuo. Theresidual water was removed by azeotropic removal with 100% ethanol. Thefinal solid was subjected to high vacuum to remove volatiles. Themixture was purified in two equal batches by preparative HPLC (C18"DELTAPAK", 60 ml/min, 48 ml fractions) using a step gradient elutionfrom 70% A/30% B to 50% A/50% B. The appropriate fractions were combined(determined by UV monitoring at λ=220 and 277 nm). Impure fractions werecombined and reprocessed in a similar fashion as described above. Atotal of 1.78 g (35% yield) of azide D-1 (Seq ID No. 46) was obtained inthis manner. ¹ H NMP, (400 MHz, CD₃ OD): δ7.02 (d, 2H), 6.69 (d, 2H),5.30 (d, 1H), 5.11 (d, 1H), 4.98 (d, 1H), 2.74 (dd, 1H), 1.13 (d, 3H).FAB-MS (Li), m/z 1081 (MH+Li)⁺.

B. Preparation of Amine of Formula (4) Compound I-1 (R^(II), R^(III) =H(Seq ID No, 1)

The purified azide compound D-1 prepared above (1.50 g) was dissolved in40 ml of methanol. 33% Aqueous acetic acid (15 ml) was added followed by0.20 g of 10% Pd-C, then the reaction vessel was flushed with N₂. Theatmosphere inside the flask was replaced with H₂ and the mixture wasstirred rapidly under an atmosphere of H₂ for 3 hours. The suspensionwas filtered through a 0.2 μm frit and the clear solution wasconcentrated to dryness in vacuo. The residue was dissolved inapproximately 20 ml of distilled water, frozen and lyophilized to obtain1.47 g (95%) of the desired amine compound (Seq ID No. 1) as a whitesolid. ¹ H NMR (400 MHz, CD₃ OD): δ7.02 (d, 2H), 6.69 (d, 2H), 5.09 (d,1H), 5.01 (d, 1H), 2.77 (dd, 1H), 1.15 (d, 3H). FAB-MS (Li), m/z 1055(MH +Li)⁺

EXAMPLE 5 ##STR11##

A. Preparation of Intermediate Benzyloxycarbonyl Compound (Seq ID No. 1)

The amine of formula (4) from Example 4 (200 mg, 0.180 mmol) andpentafluorophenyl N-benzyloxycarbonyl-3-aminopropanoate were dissolvedin 1 ml of dimethylformamide. Diisopropylethylamine (0.035 ml, 0.198mmol) was added and the mixture was stirred at ambient temperature for Ihour. The reaction mixture was diluted with 2 mls methanol and purifiedby preparative HPLC (C18 "DELTAPAK", step gradient: 70% A/30% B to 48%A/52% B, 48 ml fractions). The appropriate fractions as determined by UVabsorbance (220, 277 nm) were combined, frozen and lyophilized toproduce 100 mg (44%) of the desired intermediate. ¹ H NMR (400 MHz, CD₃OD): δ7.32 (m, 5H), 7.01 (d, 2H), 6.69 (d, 2H), 5.64 (bd, 1H), 1.18 (d,3H). FAB-MS (Li), m/z 1259 (MLi)⁺

B. Preparation of 3-aminopropanoyl Compound of formula (5); Compound I-1R^(II) =H; R^(III) =CO(CH₂)₂ NH₂ (Seq ID No. 1)

Benzyloxycarbonyl compound from Part A (94 mg, 0.075 mmol) was dissolvedin a mixture of 3 ml methanol, 1 ml of water and 0.2 ml of acetic acid.10% Pd-C (48 mg) was added and the vessel was flushed with N₂ gas. Next,the vessel was flushed with H₂ and the mixture was stirred vigorouslyunder 1 atm H₂ for 2 hours. Removal of the volatiles in vacuo gave asolid. The solid was dissolved in about 4 ml of 50% aqueousacetonitrile, frozen and lyophilized to give 80 mg (91%) of the desiredcompound of formula (5) as a whim solid. ¹ H NMR (400 MHz, CD₃ OD):δ7.01 (d, 2H), 6.69 (d, 2H), 6.67 (d, 1H), 5.10 (d, 1H), 4.99 (d, 1H),3.12 (m, 2H), 1.91 (s, 3H), 1.17 (d, 3H). FAB-MS (Li), m/z 1125 (MLi)⁺

EXAMPLE 6 ##STR12##

Preparation of N-Methylamino Compound of formula (6); CompoundI-1(R^(II) =H; R^(III) =CH3)(Seq ID No. 1)

The amine of formula (5) from Example 5 (45.6 mg, 0.135 mmol) wasdissolved in 0.5 ml of dry dimethylformamide. Iodomethane (0.021 ml,0.338 mmol) was added followed by diisopropylethylamine (0.0824 ml,0.473 mmol). After stirring at ambient temperature for 24 hours, thevolatiles were removed in vacuo and the crude product was analyzed bymass spectrometry. FAB-MS (I.,i), m/z 1068 (MLi)⁺

EXAMPLE 7 ##STR13##

A. Preparation of Intermediate Nitrile(N-Cyanomethyl) Compound I-1;R^(II) =H: R^(III) =CH₂ CN (Seq ID No. 1)

The amine compound prepared as described in Example 4 (500 mg, 0.451mmol) was dissolved in 3 ml of dry dimethylformamide. Bromoacetonitrilethat had been prepurified by passing through a small plug of magnesiumsulfate-sodium bicarbonate (0.063 ml, 0.902 mmol), was added followed bydiisopropylethylamine (0.157 ml, 0.902 mmol). The clear reaction mixturewas stirred for 12 hours and then diluted with a small volume of water.The solution was purified by preparative HPLC (C18 "DELTAPAK", stepgradient: 70% A/30% B to 47% A/53% B, 48 ml fractions). The appropriatefractions, as determined by UV absorbance at 220 and 277 nm, werepooled, frozen and lyophilized to yield 338 mg (62%) of the desiredintermediate cyanomethyl compound as a water insoluble solid. ¹ H NMR(400 MHz, CD₃ OD): δ7.01 (d, 2H), 6.69 (d, 2H), 5.12 (dd, 1H), 5.01 (dd,1H), 3.80 (s, 2H), 2.76 (dd, 1H), 1.15 (d, 3H). FAB-MS (Li), m/z 1094(MH+Li)⁺

B. Preparation of N-aminoethyl Compound of formula (7); Compound I-1;R^(II) =H; R^(III) =(CH₂)₂ NH₂ (Seq ID No. 1)

The nitrile (cyanomethyl) compound prepared above (300 mg, 0.249 mmol)was dissolved in 5.0 ml of methanol. Next, nickel (II) chloridehexahydrate (237 mg, 0.997 mmol) was added. Sodium borohydride (189 mg,4.99 mmol) was added to the solution in three portions. A blackprecipitate formed immediately and the mixture was stirred for 15minutes at ambient temperature. The heterogeneous mixture was dilutedwith about 20-40 ml of water and approximately 10-15 ml of 2N HCl wasadded. Stirring was continued for 45 minutes until the black precipitatehad dissolved and a blue-green solution remained. Purification wasaccomplished by preparative HPLC (C18 "DELTAPAK", step gradient: 70%A/30% B to 55% A/45% B, 48 ml fractions). The appropriate fractions, asdetermined by UV absorbance at 220 and 277 nm, were pooled, frozen andlyophilized to yield 180 mg (55%) of the desired product. The materialwas dissolved in 30 ml of water and passed through an ion exchangecolumn (Cl⁻ form), rinsing with distilled water. The solution was frozenand lyophilized to obtain 149 mg (94% recovery) of the desiredaminoethyl compound of formula (7) Seq ID NO. 1 as a white solid. ¹ HNMR (400 MHz, CD₃ OD): δ7.01 (d, 2H), 6.69 (d, 2H), 5.11 (dd, 1H), 5.07(dd, 1H), 1.14 (d, 3H). FAB-MS (Li), m/z 1098 (MH+Li)⁺

EXAMPLE 8 ##STR14##

A. Preparation of Intermediate Azide Compound (Seq ID No. 47)

Pneumocandin B₀ nitrile (Seq ID No. 20) (2.00 g, 1.91 mmol) wasdissolved in 24 ml of 2M LiClO₄ -diethyl ether. Triethylsilane (2.00 ml)followed by trifluoroacetic acid (1.00 ml) was added and the mixture wasrapidly stirred at ambient temperature for 6 hours. The mixture waspoured into 300 ml of water, stirred for 15 minutes and filtered. Thefilter cake was dissolved in a minimal amount of methanol and thesolvent removed in vacuo. The residual water was azeotroped with 100%ethanol and the residue was subjected to high vacuum overnight to removevolatiles to obtain a product (Seq ID No. 17) mono-reduced at thebenzylic carbon.

The crude solid from above and sodium azide (1.26 g, 19.4 mmol) wereplaced in a roundbottom flask equipped with a stirring bar and coolingbath. Trifluoroacetic acid (50 ml) was slowly added, the cooling bathwas removed and the mixture was stirred for 2 hours. It was poured into300 ml of water and filtered. The solid was dissolved in methanol,rotovaped and pumped under high vacuum to remove volatiles. The crudematerial was purified by preparative HPLC (C18 "DELTAPAK", stepgradient: 55% A/45% B to 45% A/55% B, 56 ml fractions). The appropriatefractions, as determined by UV absorbance at 220 and 277 nm, werepooled, frozen and lyophilized to yield 0.59 g (29%) of the desiredintermediate azide (Seq ID No. 47). ¹ H NMR (400 MHz, CD₃ OD): δ7.00 (d,2H), 6.69 (d, 2H), 5.34 (d, 1H), 5.07 (d, 1H), 5.00 (m, 1H), 2.88 (dd,1H), 1.17 (d, 3H). FAB-MS (Li), m/z 1036 (M-N₂ +Li)⁺

B. Preparation of Compound of Formula (8) (Seq ID. No. 48)

The purified azide from Part A (0.15 g, 0.142 mmol) was dissolved in amixture of 4 ml methanol, 1 ml water and 0.5 ml of acetic acid. 10% Pd-C(50 mg) was added to the solution. The reaction flask was flushed withN₂, then with H₂. The mixture was rapidly stirred at ambient temperaturefor 5 hours under 1 atmosphere of H₂. Subsequent filtration through a0.2 μm frit and removal of the volatiles in vacuo produced 0.124 g (80%)of the desired compound of formula (8) Compound I-2; R^(II), R^(III) =H;R^(I) =DMTD (Seq ID No. 2) as a white solid. ¹ H NMR (400 MHz, CD₃ OD):δ7.00 (d, 2H), 6.69 (d, 2H), 5.04 (d, 1H), 5.01 (m, 1H), 2.79 (dd, 1H),1.18 (d, 3H). FAB-MS (Li), m/z 1037 (MH+Li)⁺

EXAMPLE 9 ##STR15##

Preparation of Amine Compound of Formula (9) (Seq ID No. 3)

The purified azide-nitrile from Example 8, Part A (44 mg, 0.0416 mmol)was dissolved in 1.5 ml of methanol followed by CoCl₂ ·6H₂ O (59 mg,0.25 mmol). Next, NaBH₄ (8×12 mg, 2.50 mmol) was added cautiously inportions. The black, heterogeneous reaction mixture was stirred for 30minutes at ambient temperature. The reaction was quenched by addingabout 1.5 ml of 2N HCl and enough acetic acid to dissolve theprecipitate. The pale solution was diluted with 3 ml of water andpurified by preparative HPLC (C18 "ZORBAX", step gradient: 70% M30% B to60%A/40% B, 15 ml/min, 15 ml fractions). The appropriate fractions asdetermined by UV absorbance at 210 and 277 nm, were pooled, frozen andlyophilized to obtain 38 mg (72%) of the desired compound of formula (9)as a white solid. ¹ H NMR (400 MHz, CD₃ OD): δ6.99 (d, 2H), 6.70 (d,2H), 5.11 (d, 1H), 5.0 (m, 1H), 3.05 (m, 2H), 1.17 (d, 3H). FAB-MS (Li),m/z 1041 (MH+Li)⁺

EXAMPLE 10 ##STR16##

A. Preparation of Intermediate Bis-nitrile Compound (Compound I-2;R^(II) =H; R^(III) =CH₂ CN; R^(I) =DMTD) (Seq ID No. 2)

The nitrile-amine compound of Example 8 Part B (500 mg, 0.459 mmol) wasdissolved in 3 ml of dry dimethylformamide. Bromoacetonitrile that hadbeen prepurified by passing through a small plug of magnesiumsulfate-sodium bicarbonate (0,064 ml, 0.917 mmol), was added followed bydiisopropylethylamine (0.155 ml, 0.917 mmol). The reaction mixture wasstirred at ambient temperature for 18 hours. It was diluted with waterand purified by preparative HPLC (C18 "DELTAPAK", 60 ml/min, stepgradient: 70% A/30% B to 50% A/50% B, 48 ml fractions). The appropriatefractions, as determined by UV absorbance at 220 and 277 nm, werepooled, frozen and lyophilized to obtain 198 mg (36%) of the desiredCompound I-2; R^(II) =H; R^(III) =CH₂ CN ¹ H NMR (400 MHz, CD₃ OD):δ7.00 (d, 2H), 6.69 (d, 2H), 5.08 (dd, 1H), 5.01 (dd, 1H), 3.73 (s, 2H),2.79 (dd, 1H), 1.18 (d, 3H). FAB-MS (Li), m/z 1076 (MH+Li)⁺

B. Preparation of Compound of formula (10) (Seq ID No. 3)

The bis-nitrile from Part A (184 mg, 0.155 mmol) was dissolved in 3 mlof methanol. NiCl₂ ·6H₂ O (148 mg, 0.621 mmol) was dissolved in themethanol and NaBH₄ (117 mg, 3.1 mmol) was added in three portions. After5 minutes, COCl₂ ·6H₂ O (148 mg, 0.621 mmol) was added and stirred about1 minute. An additional 117 mg of NaBH₄ was added and stirring wascontinued for 15 minutes. Another 60 mg portion of NaBH₄ was added todrive the reaction to completion. The mixture was diluted with water,acidified with 2N HCl and stirred until the black precipitate dissolved.Purification by preparative HPLC (C18 "ZORBAX", 15 ml/min, stepgradient: 70% A/30% B to 55% A/45% B, 22.5 ml fractions, 220, 277 nm)gave after lyophilization a solid. The solid was dissolved in water andpassed through an ion exchange column (Cl-- form), frozen andlyophilized to give 81.1 mg (44%) of the desired compound of formula(10) (Compound I-3 (Seq ID No. 3) as a white solid. ¹ H NMR (400 MHz,CD₃ OD): δ7.00 (d, 2H), 6.70 (d, 2H), 3-3.3 (m, 6H), 1.18 (d, 3H).FAB-MS (Li), m/z 1084 (MH+Li)⁺

EXAMPLE 11-14

In operations carded out in a manner similar to that described inExample 4, the appropriate cyclopeptide natural products or modifiednatural products obtained as described hereinafter on preparation ofstarting materials are in separate operations dissolved in LiClO₄-diethyl ether and to it is added with stirring trifluoroacetic acid andtriethylsilane for 5 to 10 hours. The mixture is then poured into water,filtered, and the solid stirred with diethyl ether, then filtered andair dried to obtain cyclopeptide in which R₁ has been reduced to H.

The monoreduced compound is added to a preformed solution of HN₃ (fromNaN₃ and trifluoroacetic acid) with cooling and stirred at roomtemperature form 30 minutes to one hour and then poured into water toobtain the azide product which is recovered in the manner previouslydescribed.

The azide is hydrogenated as previously described using Pd/C as catalystand the product is recovered from the filtrate after separation of thecatalyst.

The products obtained in this manner are as follows:

    __________________________________________________________________________                                       Seq.                                       ID    Example          R.sub.1             R.sub.2                 R.sub.3                        NR.sup.II                            R.sup.III                                R.sup.I                                       No.    __________________________________________________________________________    11    H  H   CH.sub.2 CONH.sub.2                        H   H   C.sub.6 H.sub.4 OC.sub.8 H.sub.17                                       12    12    H  H   CH.sub.2 CN                        H   H   C.sub.6 H.sub.4 OC.sub.8 H.sub.17                                       13    13    H  H   CH.sub.2 CH.sub.2 NH.sub.2                        H   H   C.sub.6 H.sub.4 OC.sub.8 H.sub.17                                       14    14    H  CH.sub.3                 CH.sub.3                        H   H   C.sub.6 H.sub.4 OC.sub.8 H.sub.17                                       15    __________________________________________________________________________

EXAMPLES 15-17

In operations carded out in a manner similar to that described inExample 7, the compounds of Examples 11, 13 and 14, are dissolved indimethylformamide and added thereto are purified bromoacetonitrilefollowed by diisopropylethylamine and the mixture stirred from twelve toeighteen hours to produce a nitrile (an N-cyanomethyl) compound. Thelatter is purified by preparative HPLC.

The nitrile is dissolved in methanol and reduced chemically employingnickel (II) chloride and sodium borohydride to obtain animoethylsubstituted compound as follows:

    __________________________________________________________________________                                        Seq                                        ID    Example          R.sub.1             R.sub.2                R.sub.3                       NR.sup.II                          R.sup.III                                 R.sup.I                                        No.    __________________________________________________________________________    15    H  H  CH.sub.2 CONH.sub.2                       H  CH.sub.2 CH.sub.2 NH.sub.2                                 C.sub.10 H.sub.6 OC.sub.8 H.sub.17                                        12    16    H  H  (CH.sub.2).sub.2 NH.sub.2                       H  CH.sub.2 CH.sub.2 NH.sub.2                                 C.sub.10 H.sub.6 OC.sub.8 H.sub.17                                        14    17    H  H  CH.sub.3                       H  CH.sub.2 CH.sub.2 NH.sub.2                                 C.sub.10 H.sub.6 OC.sub.8 H.sub.17                                        15    __________________________________________________________________________

EXAMPLES 18-21

In operations carried out in a manner similar to that described inExample 1, 2 and 3, compounds having the substituents below may beprepared:

    __________________________________________________________________________                                        Seq                                        ID    Example         R.sub.1             R.sub.2                 R.sub.3                        NR.sup.II                            R.sup.III                                   R.sup.I                                        No.    __________________________________________________________________________    18   OH  CH.sub.3                 CH.sub.2 CONH.sub.2                        H   CH.sub.2 CH.sub.2 NH.sub.2                                   DMTD 7    19   OH  CH.sub.3                 CH.sub.2 CH.sub.2 NH.sub.2                        H   CH.sub.2 CH.sub.2 NH.sub.2                                   DMTD 8    20   OH  OH  CH.sub.2 CONH.sub.2                        H   CH.sub.2 CH.sub.2 NH.sub.2                                   DMTD 9    21   OH  OH  CH.sub.2 CH.sub.2 NH.sub.2                        H   CH.sub.2 CH.sub.2 NH.sub.2                                   DMTD 14    __________________________________________________________________________

EXAMPLES 22-25

In operations carried out in a manner similar to that described inExample 1, the following compounds are prepared:

    __________________________________________________________________________                                        Seq                                        ID    Example         R.sub.1             R.sub.2                R.sub.3                       NR.sub.II                          R.sub.III                                 R.sub.I                                        No.    __________________________________________________________________________    22   OH  CH.sub.3                CH.sub.3                       H  CH.sub.2 CH.sub.2 NH.sub.2                                 C.sub.6 H.sub.4 OC.sub.8 H.sub.17                                        10    23   OH  CH.sub.3                H      H  CH.sub.2 CH.sub.2 NH.sub.2                                 C.sub.6 H.sub.4 OC.sub.8 H.sub.17                                        11    24   OH  H  CH.sub.2 CH.sub.2 NH.sub.2                       H  (CH.sub.2).sub.3 NH.sub.2                                 DMTD    6    25   OH  H  CH.sub.2 CH.sub.2 NH.sub.2                       H  CH.sub.2 CH.sub.2 NH.sub.2                                 DMTD    6    __________________________________________________________________________

EXAMPLE 26 ##STR17##

The above compound is prepared in a manner similar to that described inExample 2, Part B, substituting dimethylamine for ethylenediamine toobtain a compound of M.W.=1334.43.

EXAMPLE 27 ##STR18##

The above compound is prepared in a manner similar to that in Example26, substituting piperidine for dimethylamine to obtain a compound ofM.W. 1374.

EXAMPLE 28

1000 compressed tablets each containing 500 mg of the compound offormula (2), Compound I-6 (R^(II) =H; R^(IIl) =2-aminoethyl) Seq ID No6.!, are prepared from the following formulation:

    ______________________________________    Compound              Grams    ______________________________________    Compound of Example 2 500    Starch                750    Dibasic calcium phosphate, hydrous                          5000    Calcium stearate      2.5    ______________________________________

The finely powdered ingredients are mixed well and granulated with 10percent starch paste. The granulation is dried and compressed intotablets.

EXAMPLE 29

1000 hard gelatin capsules, each containing 500 mg of the same compoundare prepared from the following formulation:

    ______________________________________    Compound           Grams    ______________________________________    Compound of Example 2                       500    Starch             250    Lactose            750    Talc               250    Calcium stearate    10    ______________________________________

A uniform mixture of the ingredients is prepared by blending and used tofill two-piece hard gelatin capsules.

EXAMPLE 30

An aerosol composition may be prepared having the following formulation:

    ______________________________________                         Per Canister    ______________________________________    Compound of Example 2  24      mg    Lecithin NF Liquid Concd.                           1.2     mg    Trichlorofluoromethane, NF                           4.026   g    Dichlorodifluoromethane, NF                           12.15   g    ______________________________________

EXAMPLE 31

250 milliliters of an injectible solution may be prepared byconventional procedures having the following formulation:

    ______________________________________    Dextrose               12.5   g    Water                  250    ml    Compound of Example 4  400    mg    ______________________________________

The ingredients are blended and thereafter sterilized for use.

PREPARATION OF STARLING MATERIALS

A-4 when R^(I) is DMTD may be produced by cultivating Zalerionarboricola ATCC 206868 in nutrient medium with mannitol as the primarysource of carbon as described in U.S. Pat. No. 5,021,341, Jun. 4, 1991.

A-7 when R^(I) is DMTD may be produced by cultivating Zalerionarboricola ATCC 20868 in nutrient medium as described in U.S. Pat. No.4,931,352, Jun. 5, 1990.

A-10 when R^(I) is linoleyl may be produced by cultivating Aspergillusnidulans NRRL 11440 in nutrient medium as described in U.S. Pat. No.4,288,549, Sep. 8, 1981.

A-11 when R^(I) is 11-methyltridecyl may be produced by cultivatingAspergillus sydowi in nutrient medium as described in J. Antibiotics XL(No. 3) p 28 (1987).

A-12 may be produced by cultivation of Zalerion arboricola ATCC 20958 innutrient medium as described in copending U.S. application Ser. No.07/630,457, filed Dec. 19, 1990, U.S. Pat. No. 5,306,708 issued Apr. 26,1994.

Compounds in which R₁ is H may be produced as described in Example 4,Part A.

Compounds in which R₃ is CH₂ CN such as A-2, A-5 and A-8 may be producedby the reaction of a compound having a carboxamide group in thecorresponding position with excess cyanuric chloride in an aproticsolvent. Molecular sieves may be employed in this reaction. Aftercompletion of the reaction, the sieves, if employed, are removed, andthe filtrate concentrated to obtain the nitrile compound as more fullydescribed in copending U.S. application Ser No. 936,434, Sep. 3, 1992.

Compounds in which R₃ is CH₂ CH₂ NH₂ such as A-3, A-6 and A-9 may beproduced by either a chemical or catalytic reduction of the nitrile. Itis conveniently carried out employing large molar excess of sodiumborohydride with cobaltous chloride as more fully described in copendingU.S. application Ser. No. 936,558, Sep. 3, 1992.

Starting materials in which R^(I) is a different group from that of thenatural product may be obtained by deacylating the lipophilic group ofthe natural product by subjecting the natural product in a nutrientmedium to a deacylating enzyme until substantial deacylation occurs,said enzyme having first been obtained by cultivating a microorganism ofthe family Pseudomondaceae or Actinoplanaceae, as described inExperentia 34, 1670 (1978) or U.S. Pat. No. 4,293,482, recovering thedeacylated cyclopeptide, and thereafter acylating the deacylatedcyclopepetide by mixing together with an appropriate active ester R^(I)COX to obtain Compound A with the desired acyl group.

    __________________________________________________________________________    SEQUENCE LISTING    (1) GENERAL INFORMATION:    (iii) NUMBER OF SEQUENCES: 60    (2) INFORMATION FOR SEQ ID NO:1:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 1:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:2:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 2:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:3:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ 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DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 10:    XaaThrXaaXaaThrXaa    15    (2) INFORMATION FOR SEQ ID NO:11:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 11:    XaaThrXaaXaaSerXaa    15    (2) INFORMATION FOR SEQ ID NO:12:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 12:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:13:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 13:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION 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Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 17:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:18:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 18:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:19:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 19:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:20:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 20:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:21:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 21:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:22:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 22:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:23:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 23:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:24:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 24:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:25:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 25:    XaaThrXaaXaaThrXaa    15    (2) INFORMATION FOR SEQ ID NO:26:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 26:    XaaThrXaaXaaSerXaa    15    (2) INFORMATION FOR SEQ ID NO:27:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 27:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:28:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 28:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:29:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 29:    XaaThrXaaXaaXaaXaa    15    (2) INFORMATION FOR SEQ ID NO:30:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 6    (B) TYPE: AMINO ACID    (C) STRANDEDNESS: Not Relevant    (D) TOPOLOGY: CIRCULAR    (ii) MOLECULE TYPE:    (A) DESCRIPTION: PEPTIDE    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 30:    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We claim:
 1. An antimicrobial composition comprising a compound selectedfrom the group consisting of ##STR19## or a pharmaceutically acceptablesalt thereof in admixture with a pharmaceutically acceptable carrier. 2.A method for controlling mycotic infections comprising administering toa mammalian subject in need of treatment, an antimycotic amount of acompound selected from the group consisting of ##STR20## or apharmaceutically acceptable salt thereof.
 3. A method for controllingPneumocystis pneumonia in immune-compromised patients comprisingadministering a therapeutically effective amount of a compound selectedfrom the group consisting of ##STR21## or a pharmaceutically acceptablesalt thereof.